Probing the histone binding requirements of the Hif1 histone chaperone, a component of the Hat1 chromatin assembly complex

Abstract

Hif1 (Hat1‐interacting factor 1) is a component of the Hat1 chromatin assembly complex. This complex contains the Hat1 histone acetyltransferase enzyme, that specifically acetylates histone H4, and acts in a pathway that deposits newly synthesized histone H3 and histone H4 onto DNA during chromatin assembly. The Hat1 complex is highly conserved, found in organisms ranging from yeast to humans, and plays a role in gene regulation and DNA repair. Hif1 binds histones and has histone chaperone activity. Evidence suggests that the Hat1 complex acts in histone deposition upstream of the Asf1 histone chaperone. Asf1 interacts with a heterodimer of histones H3 and H4 and blocks formation of the heterotetrameric form of histones H3 and H4 found in chromatin. This raises the question of whether Hif1 binds to a similar H3–H4 heterodimer or to a heterotetrameric complex. This is especially intriguing as Hif1 forms oligomers. To test this question, we have created site‐directed mutants of histone H3 and produced mutant H3–H4 complexes in bacteria via co‐expression methods. Mutations were introduced at the H3–H3 interaction surface that destabilize heterotetramer formation and produce obligate heterodimers. We are currently analyzing the ability of Hif1 to interact with native and mutant histone complexes to assess whether it prefers to bind heterodimeric or heterotetrameric histone complexes.